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PubMedDate not supplied

The effect of methoxychlor on the epididymal antioxidant system of adult rats.

Methoxychlor is widely used as a pesticide in many countries and has been shown to induce reproductive abnormalities in male rats, causing reduced fertility. The mechanism of action of methoxychlor on the male reproductive system is not clear. In the present study we investigated whether administration of methoxychlor induces oxidative stress in the epididymis and epididymal sperm of adult rats. Methoxychlor (50, 100, or 200 mg/kg body weight/day) was administered orally for 1, 4, or 7 days. The animals were killed using anesthetic ether 24 h after of the last treatment. Epididymal sperm were collected by cutting the epididymis into small pieces in Ham's F-12 medium at 35 degrees C. The body weight and weights of the testis, liver, and kidney did not show any significant changes in the methoxychlor-treated rats. The weight of the epididymis, seminal vesicles, and ventral prostate as well as epididymal sperm counts decreased after 50, 100, or 200 mg/kg/day for 7 days but remained unchanged after shorter courses of treatment. Epididymal sperm motility was decreased in a dose-dependent manner in the animals treated with methoxychlor for 4 or 7 days. The activities of the antioxidant enzymes superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase were decreased while the levels of hydrogen peroxide and lipid peroxidation were increased in the epididymal sperm as well as in the caput, corpus, and cauda epididymis after 4 or 7 days of treatment. The activities of superoxide dismutase decreased while the levels of lipid peroxidation increased in the liver but not in the kidney in all groups. Co-administration of the antioxidant vitamin E (20 mg/kg body weight/ day) to the 200 mg/kg/d methoxychlor-treated rats for 7 days prevented significant changes in the antioxidant systems in the epididymis and epididymal sperm and prevented alterations in sperm counts and motility. The results indicated that methoxychlor induces oxidative stress in the epididymis and epididymal sperm by decreasing antioxidant enzymes, possibly by inducing reactive oxygen species. In conclusion the adverse effect of methoxychlor on the male reproduction could be due to induction of oxidative stress.

Animals↗
PubMed2006

Biodegradation of methoxychlor and its metabolites by the white rot fungus Stereum hirsutum related to the inactivation of estrogenic activity.

The white rot fungus Stereum hirsutum was used to degrade methoxychlor [2,2,2-trichloro-1,1-bis(4-methoxyphenyl)ethane] in culture and the degraded products were extensively determined. The estrogenic activity of the degraded products of methoxychlor was examined using cell proliferation and pS2 gene expression assays in MCF-7 cells. S. hirsutum showed high resistance to methoxychlor 100 ppm, and the mycelial growth was fully completed within 8 days of incubation at 30 degrees C. Methoxychlor in liquid culture medium was gradually converted into 2,2-dichloro-1,1-bis(4-methoxyphenyl)ethane, 2,2-dichloro-1,1-bis(4-methoxyphenyl)ethylene, 2-chloro-1,1-bis(4-methoxyphenyl) ethane, 2-chloro-1,1-bis(4-methoxyphenyl) ethylene, and 1,1-bis(4-methoxyphenyl)ethylene, indicating that methoxychlor is dominantly degraded by dechlorination and dehydrogenation. MCF-7 cells were demonstrated to proliferate actively at the 10-5 M concentration of methoxychlor. However, cell proliferation was significantly inhibited by the incubation with methoxychlor culture media containing S. hirsutum. In addition, the expression level of pS2 mRNA was increased at the concentration (10-5 M) of methoxychlor. The reductive effect of S. hirsutum for methoxychlor was clear but not significant as in the proliferation assay.

Basidiomycota↗
PubMed2004

Evaluation of the developmental and reproductive toxicity of methoxychlor using an anuran (Xenopus tropicalis) chronic exposure model.

The chronic toxicity of methoxychlor to the South African clawed frog, Xenopus (Silurana) tropicalis, was evaluated using a life cycle approach. The chronic exposure period ranged from mid-cell blastula stage [NF (Nieuwkoop and Faber, 1994) stage 8] to 90 days of exposure, during which time the organisms generally completed metamorphosis and emerged as juvenile frogs. Methoxychlor concentrations ranged from 1 to 100 micrograms/l. Methoxychlor concentrations >10 micrograms/l caused delayed development. Organisms exposed to 10 micrograms/l methoxychlor for 30 days showed enlarged thyroid glands with follicular hyperplasia. No increase in mortality or external malformation was observed at any of the test concentrations during early embryo-larval development (NF stage 8 to NF stage 46; ca. 2 days exposure). A concentration-dependent increase in external malformations and internal abnormalities of the liver and gonads were noted after 90 days of exposure, however. Skewing of the sex ratio toward the female gender decreased ovary weight and number of oocytes, and increased oocyte immaturity and necrosis were noted at methoxychlor concentrations of 100 micrograms/l. Reductions in testis weight and sperm cell count were also detected at 100 micrograms/l methoxychlor. Results from these studies suggested that methoxychlor was capable of altering the rate of larval development, but did not adversely affect early embryo-larval development (2 days of exposure) as manifested in external malformations. Internal malformations, increases in the ratio of phenotypic females, were induced by chronic methoxychlor exposure. In addition, reproductive endpoints, most notably in the female specimens, were adversely affected by methoxychlor exposure. These studies add to the standardization and validation of a useful amphibian test methods capable of evaluating both reproductive and developmental effects of potential endocrine disrupting chemicals over a life cycle exposure.

Animals↗
PubMed2004

Effect of methoxychlor on various life stages of Xenopus laevis.

The toxicological effects of the organochlorine pesticide methoxychlor were evaluated at various life stages of the South African clawed frog, Xenopus laevis, in an effort to determine stage-specific sensitivity. A battery of four separate assays, including a short-term (4-day) early embryo-larval assay (FETAX) (NF stages 8-46 [Nieuwkoop and Faber, 1994]), 30-day hind limb development assay (NF stages 8-54), 18-day metamorphic climax assay (NF stages 58-66), and 30-day adult reproduction assay were performed. Test concentrations for the FETAX, hind limb development, metamorphic climax, and reproductive assays ranged from 0.0001-1.0 mg/l, 0.0001-0.1 mg/l, 0.0001-0.1 mg/l, and 0.001-0.1 mg/l, respectively. Results from the short-term embryo-larval assay indicated that increased embryo-lethality, malformation, and growth inhibition were not induced at </=1.0 mg/l methoxychlor (maximum soluble concentration). The 30-day hind limb development studies indicated methoxychlor exposure >/=0.01 mg/l delayed hind limb digit differentiation. Follicular hyperplasia of the thyroid glands was noted in specimens exposed to 0.1 mg/l methoxychlor. Results from the 18-day metamorphic climax assay indicated that methoxychlor inhibited the rate of tail resorption in a concentration-dependent manner. Whole body tissue triiodothyronine (T(3)) profiles showed a reduced and delayed surge during climax compared to controls. For the reproductive assessment, adult female X. laevis were super-ovulated and both female and male were then exposed to varying concentrations of methoxychlor. A concentration-dependent reduction in ovary weight and the number of viable oocytes was observed. In exposed male specimens, a concentration-dependent reduction in testis weight and sperm count was found. Methoxychlor was found to accumulate in the ovary, and to a lesser extent in the testis. Based on breeding studies in which exposed females were bred with control males and exposed males bred with control females, the frequency of amplexus, fertilization, and embryo viability was also affected by adult female methoxychlor exposure, and to a lesser extent by male exposure. Overall, these results suggested that sensitivity to methoxychlor is most dramatic during the reproductive and metamorphic phases of the life cycle and least sensitive during early embryo-larval development.

Animals↗
PubMed2005

Dietary methoxychlor exposure modulates splenic natural killer cell activity, antibody-forming cell response and phenotypic marker expression in F0 and F1 generations of Sprague Dawley rats.

Methoxychlor, a chlorinated hydrocarbon pesticide, is a persistent environmental contaminant that has been identified in human reproductive tissues. Methoxychlor has been shown to be estrogenic in both in vivo and in vitro studies. As an endocrine disrupter, it may have the potential to adversely affect endocrine, reproductive, and immune systems in animals. The present study evaluated methoxychlor's immunotoxic potential in F0 (dams) and F1 generations of Sprague Dawley rats exposed to an isoflavone-free diet containing methoxychlor at concentrations of 10, 100, and 1000 ppm. In dams, exposure to methoxychlor from gestation day 7 to postpartum day 51 (65 days total exposure) produced a significant increase in the NK activity (1000 ppm) and the percentages of T cells (1000 ppm), helper T cells (1000 ppm) and macrophages (100 and 1000 ppm). In contrast, a decrease in the numbers of splenocytes and B cells was observed at the 100 and 1000 ppm concentrations. In F1 males, exposure to methoxychlor gestationally, lactationally and through feed from postnatal day 22-64 (78 days total exposure) produced an increase in the spleen IgM antibody-forming cell response to sheep red blood cells (100 and 1000 ppm) and the activity of NK cells (1000 ppm). However, there was a decrease in the terminal body weight (1000 ppm), spleen weight (1000 ppm), thymus weight (100 and 1000 ppm), and the numbers of splenocytes (1000 ppm), B cells (100 and 1000 ppm), cytotoxic T cells (1000 ppm) and NK cells (100 and 1000 ppm). In F1 females, exposure to methoxychlor produced a decrease in the terminal body weight (1000 ppm) and the percentages of cytotoxic T cells (10, 100 and 1000 ppm). These results demonstrate that developmental and adult dietary exposure to methoxychlor modulates immune responses in Sprague Dawley rats. Immunological changes were more pronounced in the F1 generation male rats that were exposed during gestation and postpartum, when compared to the F0 and F1 generation females. Increases in antibody-forming cell response and NK cell activity, and altered spleen cell subpopulation numbers were observed in the F1 generation male rats, without similar changes to the F1 generation females.

Animals↗
PubMedDate not supplied

The diverse mechanism of action of dichlorodiphenyldichloroethylene (DDE) and methoxychlor in ovarian cells in vitro.

Dichlorodiphenyldichloroethylene (DDE), the most stable metabolite of the organochlorine insecticide dichlorodiphenyltrichloroethane (DDT), and the DDT analog methoxychlor can have adverse effects on reproduction. These chemicals have been identified as having estrogenic activity. The aim of the current study was to examine the effects of dichlorodiphenyldichloroethylene (DDE), methoxychlor, and estradiol-17 beta on steroidogenesis and FSH responsiveness in ovarian cells in vitro. Experiments were performed on a primary culture of porcine granulosa cells and a culture of Chinese hamster ovary (CHO) cells, the latter stably transfected with the FSH receptor (CHO-FSH-R). DDE (10 microM) and estradiol-17 beta (0.1 microM) but not methoxychlor (10 microM), increased proliferation of the granulosa cells. DDE (100 and 10 microM, respectively) decreased FSH-stimulated cAMP synthesis in the granulosa and CHO-FSH-R cells. DDE also decreased progesterone synthesis in the granulosa cells. Methoxychlor (10 microM) inhibited progesterone synthesis in the granulosa cells, but it did not affect the generation of cAMP in either type of cells studied. However, methoxychlor inhibited estradiol-17 beta-stimulated progesterone synthesis in the granulosa cells. We conclude that DDE primarily inhibited the generation of cAMP, while methoxychlor suppressed progesterone synthesis through a mechanism distal to cAMP generation. The present results indicate that DDE and methoxychlor are not limited to a mimicking of the endocrine effects of estradiol-17 beta in cultured ovarian cells. Therefore, a non-estrogenic component of the endocrine disrupting activities of DDE and methoxychlor must be considered in evaluating their reproductive toxicity.

Animals↗
PubMed2000

Effect of methoxychlor administration to male rats on hepatic, microsomal iodothyronine 5'-deiodinase, form I.

We previously reported that methoxychlor administration inhibits the activity of the hepatic, microsomal iodothyronine 5'-deiodinase, form I (ID-I; ). Our data further suggested that the inhibition was due to the covalent binding of a methoxychlor metabolite to a 56-kDa protein identified as ID-I (; ). This protein is 98% homologous to the thiol:protein disulfide oxidoreductase, form Q5 (ERp55;; ). Although at the time there was some controversy, most studies now suggest that ID-I is actually catalyzed by a 27-kDa selenoprotein that does not form adducts with methoxychlor (;; ). Because the 27-kDa protein is considered to be ID-I instead of ERp55, we have further examined the basis for the decreased ID-I activity observed after methoxychlor administration. Male, 150- to 200-g Sprague-Dawley rats were given methoxychlor (0-100 mg/kg/day) in corn oil by gavage for 14 days. ID-I was determined by a thyronine-specific immunoassay. Treated rats showed a significant 15% decline in total hepatic, microsomal protein at all doses. The ID-I-specific activity showed a linear decrease with increasing log doses of methoxychlor. The maximum decrease was 42% at 100 mg/kg/day. The 27-kDa protein specific content declined 37%. In rats given methoxychlor the ratios of the 27-kDa protein mRNA to the 18S ribosomal RNA declined from 2.2 +/- 0.27 x 10(-3) (controls) to 0.99 +/- 0.09 x 10(-3) (100 mg/kg/day). These data suggest that the decreased ID-I observed with chronic methoxychlor administration was due to decreased transcription or stability of the mRNA encoding the 27-kDa protein.

Animals↗
PubMed2001

Inhibition of basal and stimulated progesterone synthesis by dichlorodiphenyldichloroethylene and methoxychlor in a stable pig granulosa cell line.

The effects of the insecticide dichlorodiphenyldichloroethylene (DDE) and methoxychlor in a stable pig granulosa cell line, JC-410, were investigated. The studies of DDE and methoxychlor were conducted in combination with studies of cholera toxin, the protein kinase A activator that stimulates cAMP and progesterone synthesis and gene expression of P450 cholesterol side chain cleavage (P450scc), which converts cholesterol to pregnenolone. Administration of DDE at 3000 and 10 000 ng ml (-1) was found to decrease progesterone synthesis 0.49- and 0.25-fold, respectively, and to block the stimulatory effect of 100 ng cholera toxin ml (-1), after 24 h incubation. At 1-100 ng ml (-1), methoxychlor did not affect progesterone synthesis after 48 h incubation. However, 1000 ng methoxychlor ml (-1) decreased progesterone synthesis 0.32-fold, and both 100 and 1000 ng methoxychlor ml (-1) blocked the stimulatory effect of cholera toxin. At 3000 and 10 000 ng ml(-1), DDE decreased cAMP synthesis 0.66-and 0.36-fold, respectively. At 300, 3000 and 10 000 ng ml (-1), DDE also decreased cholera toxin-stimulated cAMP synthesis 0.84-, 0.68-, and 0.52-fold, respectively. Administration of 1-100 ng methoxychlor ml (-1) did not affect basal or cholera toxin-stimulated cAMP synthesis. Cholera toxin increased P450scc mRNA 1.4-fold after 24 h incubation, while 3000 and 10 000 ng DDE ml (-1) led to 0.39- and 0.18-fold reductions, respectively. The stimulatory effect of cholera toxin on P450scc mRNA was blocked by 3000 and 10 000 ng DDE ml(-1). Cholera toxin increased P450scc mRNA 3.48-fold after 48 h incubation, while 100 and 1000 ng methoxychlor ml (-1) increased P450scc mRNA 1.79- and 3.0-fold, respectively, and further increased the stimulatory effect of cholera toxin 6.47- and 5.44-fold, respectively. The results of the present study indicate that DDE inhibits granulosa cell steroidogenesis by affecting cAMP production and P450scc gene expression. However, methoxychlor appears to inhibit steroidogenesis by a mechanism occurring before the conversion of cholesterol into pregnenolone.

Animals↗
PubMed2002

Induction of oxidative stress in the rat testis after short-term exposure to the organochlorine pesticide methoxychlor.

Methoxychlor is one of the environmental contaminants that has been shown to induce reproductive abnormalities in male rats. The mechanism of action of methoxychlor on the male reproductive system remains unclear. In the present study we have sought to investigate whether short-term administration of methoxychlor induces oxidative stress in the testis of adult rats. Methoxychlor (50, 100, or 200 mg/kg body weight per day) was administered orally for 1, 4, or 7 days. The animals were killed using anesthetic ether on the day following the last dosing. The weights of epididymides, seminal vesicles, and ventral prostate decreased after 50, 100, or 200 mg/kg per day for 7 days but remained unchanged after 1 and 4 days of treatment. The production of superoxide anion and hydrogen peroxide increased in the animals that received methoxychlor for 4 and 7 days. The activities of the antioxidant enzymes superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase decreased, while the level of lipid peroxidation increased in the testis after 4 or 7 days of treatment. The results indicated that short-term exposure to methoxychlor induces oxidative stress in the testis by decreasing antioxidant enzymes and increasing lipid peroxidation, possibly by inducing reactive oxygen species. In conclusion, the adverse effect of methoxychlor on the male reproduction could be due to induction of oxidative stress in testis.

Administration, Oral↗
PubMed1986

Characteristics of the active oxygen in covalent binding of the pesticide methoxychlor to hepatic microsomal proteins.

This study examined the characteristics of the active oxygen species involved in generation of the reactive intermediate of methoxychlor which covalently binds to liver microsomal proteins. The possibility that the active oxygen participating in the above reaction is the superoxide anion (O2-) or a species generated from O2- was examined with the help of superoxide dismutase (SOD) and with an SOD-mimetic agent, CuDIPS [Cu2+(3,5-diisopropylsalicylic acid)2]. It was observed that, whereas CuDIPS inhibited covalent binding of methoxychlor metabolite(s), SOD did not. However, ZnDIPS [Zn2+(3,5-diisopropylsalicylic acid)2], which exhibits no SOD-mimetic activity, did not inhibit covalent binding. Furthermore, both CuDIPS and ZnDIPS had little or no effect on the formation of demethylated (polar) metabolites of methoxychlor, demonstrating that the inhibition of covalent binding by CuDIPS was not merely due to a general inhibition of the hepatic monooxygenase system. These findings suggested that O2- was involved in covalent binding, but was not accessible to SOD. Additional support for O2- involvement stems from the observation that alpha-tocopheryl acid succinate markedly inhibited covalent binding of methoxychlor. The possibility that hydrogen peroxide (H2O2) was involved in covalent binding of methoxychlor appears unlikely. Catalase had no effect on covalent binding when NADPH was the cofactor, and the use of H2O2 in place of NADPH did not yield covalent binding. Certain scavengers of hydroxyl radical (ethanol, t-butanol and benzoate) inhibited, and other known scavengers (DMSO and mannitol) did not inhibit, covalent binding. EDTA stimulated binding, desferal (desferrioxamine) exhibited no effect on binding, and diethylenetriaminepentaacetic acid (DETAPAC) inhibited binding. A possible explanation for this observation is that the Fe2+ needed for generation of X OH is much more easily obtained from Fe3+-EDTA than from Fe3+-desferal, which resists reduction. The inhibitory effect by DETAPAC may be due to chelation of another metal which is needed for the reaction. Lastly, certain scavengers of singlet oxygen inhibited covalent binding with little effect on the formation of polar metabolites of methoxychlor. In conclusion, these studies support the involvement of X OH and singlet oxygen, possibly derived from O2-, in the formation of the reactive methoxychlor intermediate.(ABSTRACT TRUNCATED AT 400 WORDS)

Benzoates↗
PubMedDate not supplied

Purified methoxychlor stimulates the reproductive tract in immature female mice.

Methoxychlor is an estrogenic pesticide currently used as a substitute for DDT. The estrogenic effects of 98% pure methoxychlor or base-washed pure methoxychlor were examined on reproductive organs of immature mice and compared to control and estradiol-treated mice. Within 24 hours of birth, neonatal mice received 14 daily intraperitoneal injections of one of the following chemicals: sesame oil only, 10.0 micrograms of 17-beta estradiol, or 0.05 mg, 0.5 mg, or 1.0 mg of 98% pure methoxychlor, or 1.0 mg base-washed 98% pure methoxychlor. Equal doses (1.0 mg) of 98% pure or base-washed methoxychlor stimulated the reproductive organs of immature females such that the effects closely resembled those seen after estradiol treatments. In comparison to control mice, these pesticide treatments also induced precocious vaginal opening, persistent vaginal cornification, increased reproductive tract weights, and epithelial hypertrophy in both the vagina and uterus. However, only estradiol treatments significantly elevated albumin levels in the uterine fluid of the immature mice while the highest methoxychlor doses significantly increased the uterine cell heights when compared to all other groups.

Albumins↗
PubMed2004

Interactions between CYP2C9 and CYP2C19 in reconstituted binary systems influence their catalytic activity: possible rationale for the inability of CYP2C19 to catalyze methoxychlor demethylation in human liver microsomes.

Previous studies in our laboratory showed that among cDNA-expressed human cytochrome P450 (P450) supersomes, CYP2C19 was the most active in methoxychlor-O-demethylation. However, based on the lack of inhibition of methoxychlor-O-demethylation by monoclonal anti-CYP2C19 antibodies in human liver microsomes (HLM), CYP2C19 did not seem to catalyze that reaction in HLM. By contrast, CYP2C9, much less active than CYP2C19 in supersomes, was the most active in HLM. The current study examines whether the lack of methoxychlor-O-demethylation by CYP2C19 in HLM was due to CYP2C19 exhibiting inferior competition for the NADPH-cytochrome P450 reductase (CPR) versus CYP2C9 and explores the interactions between CYP2C9 and CYP2C19 in a singular and binary complex of a reconstituted system. When reconstituted with CPR, cytochrome b(5), and lipid, purified CYP2C19 and CYP2C9 catalyzed methoxychlor-O-demethylation. However, whereas equimolar CPR to CYP2C9 supported maximal rates of methoxychlor demethylation and diclofenac hydroxylation, the rate of methoxychlor demethylation by CYP2C19 was not fully saturated, even with a 9-fold molar excess of CPR over CYP2C19. This behavior of CYP2C19 was also observed with S-mephenytoin as the substrate. When a binary reconstitution system was prepared by mixing CYP2C9 and CYP2C19 enzymes, methoxychlor-O-demethylation and S-mephenytoin hydroxylation by CYP2C19 were dramatically inhibited. Inhibition depended on the amount of CPR and substrate used. By contrast, in the incubation containing CYP2C9, diclofenac hydroxylation was activated by the presence of CYP2C19. These results show that interactions among P450 enzymes can modulate their catalytic rates, which depend on the substrate undergoing metabolism.

Aryl Hydrocarbon Hydroxylases↗
PubMed2002

Enantioselective metabolism of the endocrine disruptor pesticide methoxychlor by human cytochromes P450 (P450s): major differences in selective enantiomer formation by various P450 isoforms.

Methoxychlor, a currently used pesticide that in mammals elicits proestrogenic/estrogenic activity and reproductive toxicity, has been classified as a prototype endocrine disruptor. Methoxychlor is prochiral, and its metabolites 1,1,1-trichloro-2-(4-hydroxyphenyl)-2-(4-methoxyphenyl)ethane (mono-OH-M); 1,1,1-trichloro- 2-(4-methoxyphenyl)-2-(3, 4-dihydroxyphenyl)ethane (catechol-M); and 1,1,1-trichloro-2-(4-hydroxyphenyl)-2-(3, 4-dihydroxyphenyl)ethane (tris-OH-M) are chiral; whereas 1,1,1-trichloro-2, 2-bis(4-hydroxyphenyl)ethane (bis-OH-M) is achiral. These metabolites are formed during methoxychlor incubation with liver microsomes or recombinant cytochrome p450s (rp450s). Since methoxychlor-metabolite enantiomers may have different estrogenic/antiestrogenic/antiandrogenic activities than corresponding racemates, the possibility that p450s preferentially generate or use R or S enantiomers, was examined. Indeed, rCYP1A2 and r2A6 mono-demethylated methoxychlor primarily into (R)-mono-OH-M at 91 and 75%, respectively, whereas rCYP1A1, 2B6, 2C8, 2C9, 2C19, and 2D6 formed the (S)-enantiomer at 69, 66, 75, 95, 96, and 80%, respectively. However, rCYP3A4, 3A5, and 2B1(rat) weakly demethylated methoxychlor without enantioselectivity. Human liver microsomes generated (S)-mono-OH-M (77-87%), suggesting that CYP1A2 and 2A6 display only minor catalytic contribution. P450 inhibitors demonstrated that CYP2C9 and possibly 2C19 are major hepatic catalysts forming (S)-mono-OH-M, and CYP1A2 is primarily involved in forming the (R)-mono-OH-M. Demethylation rate of (S)-mono-OH-M versus (R)-mono-OH-M forming achiral bis-OH-M by rCYP1A2 was 97/3, compared with 15/85 and 17/83 for rCYP2C9 and 2C19, respectively, indicating opposite substrate enantioselectivity of rCYP1A2 versus 2C9 and 2C19. Also, rCYP1A2 preferentially O-demethylated (R)-catechol-M into (R)-tris-OH-M (at 80%), contrasting r2C9 and r2C19 that yielded (S)-tris-OH-M at 80 and 77%, respectively. Ortho-hydroxylation of mono-OH-M into catechol-M and bis-OH-M into tris-OH-M was primarily by 3A4 and was not enantioselective. In conclusion, enantiomeric abundance of methoxychlor metabolites depends on the relative catalytic activity of the hepatic p450 isoforms.

Cytochrome P-450 Enzyme System↗
PubMed1982

Enhanced biodegradation of methoxychlor in soil under sequential environmental conditions.

Ring-U-[14C]methoxychlor [1,1-bis(p-methoxyphenyl)-2,2,2-trichloroethane] was incubated in soil under aerobic and anaerobic conditions. Primary degradation of methoxychlor occurred under anaerobic conditions, but not under aerobic conditions, after 3 months of incubation. Analysis of soil extracts, using gas chromatography, demonstrated that only 10% of the compound remained at initial concentrations of 10 and 100 ppm (wt/wt) of methoxychlor. Evidence is presented that a dechlorination reaction was responsible for primary degradation of methoxychlor. Analysis of soils treated with 100 ppm of methoxychlor in the presence of 2% HgCl2 showed that 100% of the compound remained after 3 months, indicating that degradation in the unpoisoned flasks was biologically mediated. Methanogenic organisms, however, are probably not involved, as strong inhibition of methane production was observed in all soils treated with methoxychlor. During the 3-month incubation period, little or no evaluation of 14CO2 or 14CH4 occurred under either aerobic or anaerobic conditions. Cometabolic processes may be responsible for the extensive molecular changes which occurred with methoxychlor because the rate of its disappearance from soil was observed to level off after exhaustion of soil organic matter. After this incubation period, soils previously incubated under anaerobic conditions were converted to aerobic conditions. The rates of 14CO2 evolution from soils exposed to anaerobic and aerobic sequences of environments ranged from 10- to 70-fold greater than that observed for soils exposed solely to an aerobic environment.

Aerobiosis↗
PubMed2002

Methoxychlor stimulates the mouse lactoferrin gene promoter through a GC-rich element.

The lactoferrin gene in the mouse uterus is a target gene for natural estrogens and xenoestrogens. One of the xenoestrogens is methyoxychlor, an insecticide that displays both estrogenic and antiandrogenic activities. Recently, methyoxychlor was found to stimulate lactoferrin gene expression in the uterus of an estrogen receptor null mouse. The present study is designed to uncover the methoxychlor response region in the mouse lactoferrin gene promoter. A series of different lengths of the mouse lactoferrin gene 5' flanking region were linked to a chloramphenicol acetyltransferase (CAT) reporter construct and transfected into human endometrial carcinoma HEC-1B cells, an estrogen receptor null cell line, in order to examine the methoxychlor response. The transfected cells were treated with methoxychlor or the metabolite of methoxychlor, HPTE, and the CAT reporter activities were measured. Constructs that contain a mouse lactoferrin 5' region longer than 100 bp were activated more than twofold by both methoxychlor and HPTE. The activation of the CAT reporter by the chemicals was dose dependent and reached saturation. Additional deletion mutants within the 100-bp region were tested, and a GC-rich sequence (GC-II) that we have previously characterized as an epidermal growth factor (EGF) response element was identified to be the region for the methoxychlor response. GC-II binds Sp1, Sp3, and IKLF transcription factors, collaborates with the AP1/CREB binding element, and confers the EGF response. Whether the effect of methoxychlor requires the AP1/CREB binding element has yet to be established; however, the present finding provides an alternative signaling pathway for the xenoestrogens.

Animals↗
PubMed1980

Carcinogenicity and toxicity of methoxychlor.

Methoxychlor is carcinogenic for the liver of C3H and BALB/c mice and Osborne-Mendel rats, and possibly for the liver of dogs. Methoxychlor is also carcinogenic for the testis of BALB/c male mice, bone of B6C3F1 female mice, and the ovary of Osborne-Mendel female rats. The incidences of carcinomas of the liver were increased in C3H male mice and BALB/c male and female mice fed methoxychlor. There also was an increase in malignant neoplasms at all sites in BALB/c male and female mice. C3H and BALB/c male mice were more susceptible to the carcinogenic effects of methoxychlor than were female mice. BALB/c mice were more susceptible than C3H mice. Osborne-Mendel male and female rats developed significant incidences of carcinomas of the liver. The incidence of sarcomas of the spleen and abdomen, mostly hemangiosarcomas, was increased in male rats. Neoplasms of the pituitary, adrenals, and mammary gland were also increased in methoxychlor-treated female rats. Miniature swine given methoxychlor developed chronic renal disease in relatively short periods of time. There also was hyperplasia of the mammary gland and uterus, suggesting an estrogen-like effect on those organs. Methoxychlor applied to the skin of rabbits caused a dose-related atrophy of the testes, as well as chronic renal disease. Atrophy of the testes and chronic renal disease could not be evaluated in mice and rats because of insufficient data.

Animals↗
PubMed1999

Effects of endocrine-disrupting contaminants on amphibian oogenesis: methoxychlor inhibits progesterone-induced maturation of Xenopus laevis oocytes in vitro.

There is currently little evidence of pollution-induced endocrine dysfunction in amphibia, in spite of widespread concern over global declines in this ecologically diverse group. Data regarding the potential effects of endocrine-disrupting contaminants (EDCs) on reproductive function in amphibia are particularly lacking. We hypothesized that estrogenic EDCs may disrupt progesterone-induced oocyte maturation in the adult amphibian ovary, and tested this with an in vitro germinal vesicle breakdown assay using defolliculated oocytes from the African clawed frog, Xenopus laevis. While a variety of natural and synthetic estrogens and xenoestrogens were inactive in this system, the proestrogenic pesticide methoxychlor was a surprisingly potent inhibitor of progesterone-induced oocyte maturation (median inhibitive concentration, 72 nM). This inhibitory activity was specific to methoxychlor, rather than to its estrogenic contaminants or metabolites, and was not antagonized by the estrogen receptor antagonist ICI 182,780, suggesting that this activity is not estrogenic per se. The inhibitory activity of methoxychlor was dose dependent, reversible, and early acting. However, washout was unable to reverse the effect of short methoxychlor exposure, and methoxychlor did not competitively displace [3H]progesterone from a specific binding site in the oocyte plasma membrane. Therefore, methoxychlor may exert its action not directly at the site of progesterone action, but downstream on early events in maturational signaling, although the precise mechanism of action is unclear. The activity of methoxychlor in this system indicates that xenobiotics may exert endocrine-disrupting effects through interference with progestin-regulated processes and through mechanisms other than receptor antagonism.

Animals↗
PubMed1983

Covalent binding of [14C]methoxychlor metabolite(s) to rat liver microsomal components.

[14C]Methoxychlor was incubated with NADPH-fortified liver microsomes from male rats, and covalent binding to microsomal components was determined. The binding process was markedly enhanced when microsomes from phenobarbital-treated rats were employed. However, when microsomes from methylcholanthrene-treated rats were used the level of binding was not significantly affected. Incubation in the presence of glutathione, cysteine, or ascorbate markedly diminished binding. Metyrapone and SKF 525-A, inhibitors of hepatic cytochrome P-450-linked monooxygenase activity, inhibited the binding. Also, ethylmorphine and hexobarbital, alternate substrates of the monooxygenase system, inhibited binding. There was no binding to microsomal components in the absence of NADPH or oxygen. TCPO (1,1,1-trichloropropane-2,3-oxide), an inhibitor of epoxide hydrase activity, failed to enhance the binding process. However, N,N'-diphenyl-p-phenylenediamine (NDP) and n-propyl gallate (PG), both free radical scavengers, decreased binding at micromolar concentrations without altering the extent of formation of polar [14C]methoxychlor metabolites. It was concluded that methoxychlor undergoes a hepatic microsomal monooxygenase(s)-mediated activation and that the resultant reactive metabolites (possibly free radicals) bind covalently to microsomal components. By contrast, the binding resulting from the incubation of an impure mixture of polar [14C]methoxychlor metabolites with liver microsomes did not require NADPH and O2 and was not affected by NDP, Pg, ascorbate, or heat-treatment of microsomes. This finding suggested that the binding subsequent to the initial metabolic activation of methoxychlor does not require further enzymatic transformation. However, whether the binding with metabolites represents the same chemical species as the binding with [14C]methoxychlor remains to be established.

Animals↗